Modification of pertussis toxin

A method of preparing a pertussis holotoxin having a modified biological activity, which comprises:

• (A) identifying at least one site in pertussis holotoxin that interacts with a molecule that is capable of forming a complex with the holotoxin and which molecule is a substrate which is a GTP-binding protein, an .alpha.-subunit of a GTP-binding protein, or a (C-terminal fragment of an .alpha.-subunit of a GTP-binding protein and which site contributes to toxicity, cell binding or enzymatic activity of pertussis holotoxin by:
• (a) forming a crystalline complex between at least a portion of pertussis holotoxin which is pertussis holotoxin, an analog thereof, a subunit, a position of a subunit or a combination of subunits and said molecule, including exposing crystals of the at least a portion of the holotoxin to the molecule, under conditions to effect formation of the crystalline complex without substantial disruption of the crystals;
• (b) determining the three-dimensional structure of the complex by X-ray crystallography;
• and
• (c) analysing the structure to identify the at least one interacting site;
• and
• (B) modifying said identified at least one site to alter the toxicity, cell binding or enzymatic activity of said pertussis holotoxin.

A method of preparing a pertussis holotoxin having a modified biological activity, which comprises:

• (A) identifying at least one site in pertussis holotoxin that interacts with molecule that is capable of forming a complex with the holotoxin and which molecule is a substrate which is NM or a substantially non-hydrolysable analog of NAD and which site contributes to toxicity, cell binding or enzymatic activity of pertussis holotoxin by:
• (a) forming a crystalline complex between at least a portion of pertussis holotoxin which is pertussis holotoxin, an analog thereof, a subunit, a portion of a subunit or a combination of subunits and said molecule, including exposing crystals of the at least a portion or the holotoxin to the molecule, under conditions to effect formation of the crystalline complex without substantial disruption of the crystals;
• (b) determining the three-dimensional structure of the complex by X-ray crystallography;
• (c) analyzing the structure to identify the at least one interacting site;
• and
• (B) modifying said identified at least one site to alter the toxicity, cell binding or enzymatic activity of said pertussis holotoxin.